The Kazal inhibitors are serine protease inhibitors (serpins) with sequence similarity to the pancreatic secretory trypsin inhibitors (PSTIs) which appear to be present in all vertebrates. Some examples of Kazal-type inhibitors include the acrosin-trypsin inhibitor (ATI) and PEC-60 (peptide with N-terminal Glu and C-terminal Cys) isolated from porcine intestine.
PSTI is secreted from pancreatic acinar cells into pancreatic juice. Its physiological role is thought to be the prevention of trypsin-catalyzed premature activation of zymogens within the pancreas and the pancreatic duct. Since PSTI is also found in serum and in various normal and malignant tissues, it may have additional roles. Horii, A. et al. cloned and characterized the PSTI gene from human (1987; Biochem. Biophys. Res. Commun. 149:635-641) and from rat (1989; Biochem. Biophys. Res. Commun. 162:151-159). Both rat and human PSTI are translated as precursor proteins of 79 amino acids with 23 amino acid signal sequences. PSTI contains six cysteine residues, four of which are contained in the Kazal inhibitor consensus sequence motif located at position 39 to 67 of SEQ ID NO: 1.
Tumor-associated trypsin inhibitor (TATI), which is identical to PSTI, is synthesized by several tumors and cell lines. It was initially detected in the urine of patients with ovarian cancer (Stenman, U. H. et al. (1982) Int. J. Cancer 30:53-57; Huhtala, M. L. et al. (1982 ) J. Biol. Chem. 257:13713-13716). TATI is also produced by the mucosa of the gastrointestinal tract where it may protect the mucosal cells from proteolytic breakdown. Elevated levels of TATI in serum and urine occur particularly with mucinous ovarian cancer and may occur in nonmalignant diseases, e.g., pancreatitis, severe infections, and tissue destruction (Stenman, U. H. et al. (1991) Scand. J. Clin. Lab. Invest. 51(suppl. 207):5-9).
Analysis of the amino acid sequences of Kazal-type inhibitors shows some regions of similarity with other serpins, but the highly conserved cysteine residues at positions 32, 39, 46, 58, 61 and 75 (human PSTI precursor sequence numbering) solidify their group association and provide some alignment with the beta subunits of the glycoprotein hormones, chorionic gonadotropin, luteinizing hormone, follicle stimulating hormone and thyroid stimulating hormone. The reported active site, P1-P', for the Kazal type inhibitors resides in the residues at positions 41 and 42 of human PSTI precursor and may be responsible for the specificity of the inhibitor (Liepinsh, E. et al. (1994) J. Mol. Biol. 239:137-53; Fink, E et al. (1990) FEBS Lett. 270:222-4).
The precursor of the PSTI homolog PEC-60 is 86 amino acids long and has a 26 amino acid signal sequence. PEC-60 is a secreted protein with about 41% similarity to PSTI, but it does not inhibit trypsin nor does it affect basal plasma insulin levels. The known roles of PEC-60 are the in vivo inhibition of glucose-induced secretion of insulin and the regulation of cyclic adenosine monophosphate (cAMP) levels through dose-dependent reduction of cAMP formation. High levels of PEC-60 have been recorded in bone marrow and peripheral blood, and moderate levels in spleen. The results of radioimmunoassays show that pig PEC-60 is formed, stored and secreted from monocytes. High levels of PEC-60 are known to be immunoreactive in catecholamine neurons (Laazik, J. and R. Sillard (1993) Biochem. Biophys. Res. Commun. 197:849-52; Metsis, M. et al. (1992) J. Biol. Chem. 267:19829-32; and Ahren, B. et al. (1992) Pancreas 7:443-6).
PEC-60 has been shown to reduce dopamine utilization in the caudate nucleus which suggests that PEC-60 has a role in activities or diseases of the central nervous system (Laasik, supra). A Kazal-type inhibitor with amino acid sequence similarity to PEC-60 was recently identified in rat bile/pancreatic juice and has been shown to stimulate the release of cholecystokinin (Agerberth, B. et al. (1989) Proc. Nat. Acad. Sci. 86:8590-4). ATI, which is synthesized in the testes and has been isolated from seminal fluid of several species, may play a role in regulating the effects of glycoprotein hormones. Given the region of sequence similarity between the Kazal-type inhibitors and the beta subunits, inhibition may depend on blocking the glycoprotein hormone receptors in the reproductive tissues (Perry, A. C. et al. (1993) Biochim. Biophys. Acta 1172:159-60; Fink et al., supra). PSTI is expressed in internal organs such as liver and pancreas, in response to inflammatory cytokines. Expression was reported in tissues from patients with liver, pancreatic, gastric or colonic cancers, but it has also been associated with Crohn's disease, chronic hepatitis and cirrhosis (Ohmachi, Y. et al. (1994) J. Hepatol. 21:1012-6, Int. J. Pancreatol. 15:65-73, and (1994) Int. J. Cancer 57:139; Halme, L. et al. (1993) Scand. J. Clin. Lab. Invest. 53:359-66). Similarly, the level of tumor-associated trypsin inhibitor detected in ovarian cancer serves as a marker for the grade or stage of the tumor (Medl, M. (1995) Br. J. Cancer 71:1051-4; Pectasides, D. (1994) Am. J. Clin. Oncol. 17:307-12).
Discovery of a novel human Kazal-type inhibitor and the polynucleotides which encode it satisfies a need in the art by providing new compositions useful in diagnosing and treating disorders of the digestive system, including inflammation and cancer.